Summary of Work: A system is now established to determine effects of a site-specific double-strand break (DSB) in dispensable DNA in yeast. A long-lived DSB can lead to extended G2 arrest and eventual lethality. The SIR4, SIR2 and SIR3 genes are components in the DSB signaling response. These genes affect transcriptional silencing, chromatin organization, aging and DSB endjoining. SIR genes are important in the adaptation and lethality resulting from the DSB. Sir+ cells rapidly adapt to the DSB and, following a brief G2 arrest, give rise to microcolonies. Sir mutants exhibit long G2 arrest. The mutant cells slowly adapt to the break, divide and re-arrest in the next G2. These SIR-mediated effects are indirect since adaptation to the G2 checkpoint was unaffected and viability was restored in sir? strains deleted for the silent mating-type loci HML and HMR. Since deletion of RAD50 had no effect on cell viability, toleration of the DSB is not through an end-joining pathway requiring the direct participation of Sir genes. Thus, SIR4 has no direct effect on DSB repair and that there are mating type controlled genes affecting tolerance. The toleration of ionizing radiation damage in yeast requires many DNA repair and checkpoint genes, most having human homologues. A genome-wide screen of diploid mutants homozygous for deletions of 3670 non-essential genes revealed 107 new loci that influence gamma-ray sensitivity. Many appear to affect replication, recombination and checkpoint functions. Nearly 90% were sensitive to other agents and most new genes could be assigned to the following functional groups: chromatin remodeling, chromosome segregation, nuclear pore formation, transcription, golgi/vacuolar activities, ubiquitin-mediated protein degradation, cytokinesis, mitochondrial activity, and cell wall maintenance. Over 50% share homology with human genes, including 16 implicated in cancer, suggesting a large set of newly identified human genes that may have related roles in toleration of radiation damage.